1. Field of the Invention
This invention is related to and has among its objects the provision of novel cell culture media and novel methods of culturing cells. It is a particular object of the invention that the cell culture media be free of non-human proteins. Further objects of the invention will be evident from the following Description wherein parts and percentages are by weight unless otherwise specified.
2. Description of the Prior Art
Human and other mammalian cells are generally grown in a medium containing defined nutrients and a supplement, usually animal blood proteins, particularly fetal calf serum. The use of animal serum proteins requires extensive purification of the product grown in the medium, especially where the cell-induced product is for human use such as in the case of a hybridoma antibody preparation or interferon production. It is, thus, preferred to utilize homologous proteins of human origin as a supplement in these cell cultures if such proteins may be found.
Keay in Biotechnology and Bioengineering, 18, 363, (1976) describes the use of an autoclavable, low cost, serum-free medium consisting of minimum essential nutrients, bactopeptones, and insulin. The author demonstrated that the cell growth of five different cell lines was about the same as those propagated in conventional systems containing animal blood proteins.
The invention disclosed in U.S. Pat. No. 4,198,479 (hereinafter '479) provides an improved process for the growth of lymphoblastoid cells, such as, for example, Namalva cells, in a medium supplemented with human albumin. The invention further provides for the production of interferon by viral infection of these lymphoblastoid cells in a medium also supplemented with human albumin.
One problem with the use of human albumin in cell culture media is that albumin, a therapeutically useful product, is consumed. Since the supply of this raw material is limited, it is disadvantageous to direct a portion of this material to a non-therapeutic use. Furthermore, human albumin is expensive and, thus, apparently would increase the cost of the cell culturing activity and the products made thereby.
Lambert et al in J. Cell Sci., 35, 381-392, 1979, disclose a "polypeptide" with a growth factor activity in the less than 10,000 Daltons fraction (NSILA method of purification of calf serum, Rinderknecht et al, Proc. natn. Acad. Sci. USA, 73, 2365-2369, 1976). This fraction was used in the absence of whole serum in a growth medium. In attempts by the authors to purify the growth factor it was found that growth factor activity could be recovered, particularly in fraction IV, in fractions of serum prepared by the cold ethanol method of fractionation (Cohn et al, J. Am. Chem. Soc., 68, 459-475, 1956).
Recently, Maciag et al disclosed (Science, 1981, Vol. 211, 1452-1454) a cell culture medium consisting of Medium 199 containing epidermal growth factor, tri-iodothyronine, hydrocortisone, Cohn Fraction IV, insulin, transferrin, bovine brain extract, and trace elements. The Cohn Fraction IV, a potent source of insulin-like growth factors, was prepared by extraction in acetic acid for 2 hours followed by gentle boiling for 30 minutes, conditions known to result in protein denaturation. The extract was centrifuged, neutralized, dialyzed, and lyophilized.